The methionine sulfoxide reductase (Msr) system has been shown to play an important role in protecting cells against oxidative damage. This family of enzymes can repair damage to proteins resulting from the oxidation of methionine residues to methionine sulfoxide, caused by reactive oxygen species. Previous genetic studies in animals have shown that increased levels of MsrA, an important member of the Msr family, can protect cells against oxidative damage and increase life span. We have developed a high throughput screening (HTS) compatible assay to search for both activators and inhibitors of MsrA, based on the oxidation of NADPH. This project is being done in collaboration with the Scripps Florida Research Institute, which has a high throughput screening facility and an extensive chemical library. The assay involves a coupled reaction that uses reduced thioredoxin as the reducing agent for the MsrA reaction. Previous studies had shown that MsrA has a broad specificity and can reduce a variety of methyl sulfoxide compounds, including DMSO. Since the chemicals in the library are dissolved in DMSO, the assay uses DMSO as the substrate for the MsrA enzyme. The assay can also be used to detect inhibitors of the reaction which could be used in research studies. A specific activator of MsrA could have important therapeutic value for diseases that involve oxidative damage, especially age-related diseases. PUBLIC HEALTH RELEVANCE: Oxidative damage, resulting from the production of reactive oxygen species (ROS) within cells, is believed to be a major factor in age-related diseases and the aging process. The primary goal of these studies is to develop a high throughput screening assay to look for compounds that could increase the activity of MsrA, since it is known that increased levels of MsrA can protect animals against oxidative damage and increase their life span. A specific activator of MsrA in tissues could have important therapeutic value for the treatment of diseases that involve oxidative damage, especially age-related diseases.